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• Protein synthesis “Translation”
The letters of the nucleic acid is translated into amino acids.
“from nucleotide language to amino acid language”
• Genes specify the amino acids sequence in proteins.
Genetic code: the relation between the sequence of bases in
DNA (or it’s transcript RNA) and the sequences of amino acid
in protein.
• Features of Genetic Code
- Coding ratio (3 base-code)
- We have 4 bases and 20 amino acid:
Single-base code = 4
Two-base code = 4 * 4 = 16
Three-base code = 4 * 4 * 4 = 64
A. Three-base code
B. More than code can specify one amino acid
An amino acid is coded by three bases called “codon” and these
condones:
- Non-overlapping
- The sequence of bases is read sequentially from a fixed starting point.
- There are no commas between these triplets.
• The genetic code is specific
Specific codon always codes for the same amino acid
• Redundant
- For a given amino acid may have more than one codon for it.
- Codons that specify the amino acid are called “synonyms” most of
them differ only in the last base of the triplet
UUU
UUC
phe
• Universal
The genetic code almost universal in the whole of prokaryotic, plant,
animal kingdoms, the same codon used for the same amino acid.
- With few exceptions: like in the mitochondria
Codon
Common code
Mitochondrial code
AUC
Ile
Met
AGA
Arg
STOP
AGG
Arg
STOP
UGA
STOP
Trp
START
AUG
STOP
UAA , UAG UGA
U
C
A
G
U
C
A Third
G letter
U
C
A
G
U
C
A
G
of
codon
• Consequences of altering the nucleotide sequence “mutation”
A. Base substitution “Point mutation”
- Changing a single nucleotide base on the m-RNA chain, and this can
lead to:
1. Silent mutation
The codon containing the changed base codes for the same amino acid
UCA
silent
Serine
2.
UCU
Serine
Missense
The change results in a new different amino acid
UCA
missense
Serine
CCA
Proline
3. Non-sense mutation
The change leads to premature termination if the codon containing the
changed base become a termination codon.
UCA
Serine
non-sense
UAA
STOP codon
B. Base Deletion or Insertion
1. Frame shift mutation
Insertion or deletion of one or two bases will alter the
reading frame and this cause extensive change in the
translated protein  absolutely different protein
2. Insertion or deletion of one codon “3 nucleotides”
This lead to addition of new amino acids (if three bases were
inserted), or to deletion of one amino acid (if three bases
were deleted).
The reading frame in this case is not changed and the
produced protein is not extensively changed.
Missense Mutation
Non-Sense Mutation
• The Major Participants in Translation
A large number of components are required for the synthesis of
polypeptides
1. Amino acids: absence of 1 amino acid  termination of the
polypeptide at that amino acid
2. m-RNA: act as template for protein synthesis.
3. t-RNA: adaptors
4. Functional Ribosomes: protein synthesis machine.
5. Energy sources
6. Translation factors
7. Enzymes
- The translation takes place in the cytosol
• t-RNA
- At least one specific t-RNA is required for each amino acid. In
human there are 50 types of tRNA and in prokaryotes there are
30 – 40 tRNA
- 20 amino acid  more than tRNA type for a given amino acid
- tRNA has uncommon and modified bases (Inosine, Pseudouracil, … )
- All tRNA types have a common structure
• tRNA structure
- Two functional parts
A. Acceptor stem (amino acid
attachment site)
3’-terminus of tRNA has always
the sequence 5’ … CCA-OH 3’
A. Anti codon
Three base nucleotide sequence.
That recognize a specific codon
on the mRNA and they are
complementary and anti parallel,
the codon specifies the amino
acid that will be inserted into the
growing polypeptide.
tRNA Structure
• Codon Recognition by tRNA
- Recognition of a codon in the mRNA is
accomplished by anti codon sequence of the
tRNA
- Some tRNA can recognize more than codon
- Anti codon + codon binding follows the
complementary and anti parallel binding
• Wobble hypothesis
- The base at the 5’- end of anti codon is not
spatially defined and this allows nontraditional base pairing with the 3’- base of
the codon.
- The result of wobbling is that there need
not be 61 tRNA types to read the 61
codons that code for the amino acids
Anti codon
Codon
Anti codon
3’… UAC …5’
5’ …AUG …3’
5’ …CAU …3’
Wobble
position
•Coupling of tRNA to amino acids
- Amino acids are covalently
attached to OH group of the
ribose sugar of the adenosine
residue at the 3’- end of tRNA.
- Each aminoacyl tRNA
synthestase recognizes a specific
amino acid and the tRNAs that
correspond to that amino acid.
- These enzymes are highly
specific
tRNA – amino acid = activated
amino acid or charged tRNA.
• Ribosomes
Machines for protein synthesis.
- rRNA – protein complex
- Major cell constituents, an
E. coli contains 15000
ribosomes forming 25% of the
dried cell
- In eukaryotic cell the
ribosomes either free in the
cytosol or in close association
with endoplasmic reticulum
(ER)
- Mitochondria contains their own
set of ribosomes.
• Ribosomal proteins
- These proteins play
important roles in
the structure and
function of the
ribosome.
• The Mechanism of Translation
-The pathway of protein synthesis is called translation.
-Because the language of nucleotides of the mRNA is
transcripted into amino acid language.
- The mRNA is translated in 5’  3’ direction producing
polypeptide from it’s amino terminal end to its carboxylic
terminus.
- One prokaryotic mRNA can code for different polypeptide
types (poly cistronic). Because m-RNA contains different coding
regions with different initiators.
5’
3’
AUG
UAA
Code for protein A
AUG
UAG
Code for protein B
Each eukaryotic mRNA code only for one polypeptide (mono cistronic)
•Steps in protein synthesis
The small ribosomal subunit
A. Initiation
(Shine-Dalgarno
sequence)
Formyl group is added to the
charged tRNA met by the
enzyme transformylase (formyl
THF is the source)
Will be Met in
eukaryotes
The formyl group will be
removed during the elongation
GTP
The release of IF3
increase the affinity
to the large
ribosomal subunit
The Met amino acid will be
cleaved from the polypeptide.
Specifies the next a.a
• The binding of mRNA to 30 S ribosomal subunit
The 16S rRNA has a nucleotide sequence near it’s 3’ – end that
complementary to Shine-Dalgarno sequence (nucleotide bases 5’ –
UAAGGAGG – 3’ located 6 – 10 bases up stream to the AUG codon on
the mRNA)
- The mRNA 5’- end and 3’- end of rRNA (in the 30S ribosomal subunit)
can form complementary base pair and this can facilitate the binding of
the mRNA to 30S ribosomal unit.
B. Elongation
The addition of
a.a to the
carboxyl end of
the growing
polypeptide chain.
The delivery
of a.a - tRNA
to A site
GTP
Peptidyl
transferase
(integral
part of 50S
subunit)
Elongation
This process will be
repeated until a
termination codon is
reached.
By each cycle the
polypeptide has grown by
one residue and consumed
two GTP.
GTP
Translocation: Moves
by 3 nucleotides
C. Termination
Termination
codons
RF1 recognizes UAA
and UAG
UAA
RF2 recognizes UAA
and UAG
RF3 is GTPase
(stimulate the release
process via GTP binding
and hydrolysis)
UAG
UGA
GTP
•Polyribosomes (polysomes)
Many ribosomes can simultaneously translate one mRNA.
• Energetic of translation
- The energy cost for protein synthesis is high.
- The total energy required for synthesizing a protein of N residues.
2N ATPs are required to charge tRNAs
1 GTP is needed for initiation.
N –1 GTPs are needed to form N –1 peptide bonds
N –1 GTPs are needed to form N –1 translocation steps
1 GTP is needed for termination
So the total energy:
2N+1 + N-1 + N-1 + 1 = 4 N
• Post translational modification. ”The final stage of protein synthesis”
Folding and covalent modification.
The produced protein may fold to form the 3° structure and may associate with
other subunits.
The covalent modification involve:
Phosphorylation, Glycosylation, Hydroxylation
Trimming
Protein synthesis “Translation”
The End
GOOD LUCK
•Coupling of tRNA to amino
acids
- Amino acids are covalently
attached to OH group of the
ribose sugar of the adenosine
residue at the 3’- end of
tRNA.
- Each aminoacyl tRNA
synthestase recognizes a
specific amino acid and the
tRNAs that correspond to that
amino acid.
- These enzymes are highly
specific
tRNA – amino acid = activated
amino acid or charged tRNA.
Activation of
the amino acid
AminoacyltRNA
synthestase
Adding the amino
acid to the specific
tRNA
Formation of
ester bond
The small ribosomal subunit
• Steps in protein synthesis
A.
Initiation
Formyl group is added to the
charged tRNA met by the enzyme
transformylase (formyl THF is the
source)
Will be Met
in
eukaryotes
(Shine-Dalgarno
sequence)
The release of IF3
increase the affinity
to the large ribosomal
subunit
The formyl group will be
removed during the
elongation
The Met amino acid will be
cleaved from the polypeptide.
Specifies the next a,a
The delivery of
a.a - tRNA to A
site
By each cycle the
polypeptide has grown by
one residue and consumed
two GTP.
This process will be
repeated until a termination
codon is reached.
Peptidyl
transferase
(integral
part of 50S
subunit)
Moves with 3
nucleotides
Translocation
C.
Termination
RF1 recognizes UAA and UAG
UAA
RF2 recognizes UAA and UAG
UAG
RF3 is GTPase (stimulate the
release process via GTP
binding and hydrolysis)
UGA